*Target (protein/gene name): Inositol phosphate phosphatase sopB *NCBI Gene # or RefSeq#: NC_003197.1 *Protein ID (NP or XP #) or Wolbachia#: 4DID -- (need the NP or XP number here - put the PDB number down further 082413 - Dr. B) *Organism (including strain): Salmonella enterica subsp. enterica serovar Typhimurium Etiologic Risk Group (see link below): group 2 *Background/Disease Information (sort of like the Intro to your Mini Research Write up): The salmonella enterica serovar Typhi organism is responsible for typhoid fever. The bacteria works by invading the surface of the intestine, and spreading to deeper tissues (including that of the liver, spleen, and bone marrow). It causes 22 million cases of typhoid around the world and 200,000 deaths. It is most common in underdeveloped areas that do not have proper sanitation techniques, as it is transmitted through drinking water or eating food that is contaminated with the feces of an infected person. Essentiality of this protein: SopB is an effector protein with phosphoinositide phosphatase activity. It has a distinct GTPase binding domain that interacts with the host Cdc42 cell to regulate critical events in eukaryotic cytoskeleton organization and membrane trafficking. The SopB protein mimics a host guanine nucleotide dissociation inhibitor both structurally and functionally by contacting key residues in the regulatory regions of Cdc42, which slows nucleotide exchange in the Cdc42 cell. It also prevents lysosomal degradation of the bacteria Complex of proteins: May be complexed with the host Cdc42 cell. Druggable Target:http://tdrtargets.org/drugs/view?mol_id=558832 *EC#: 3.1.3.36 Link to BRENDA EC# page:
Fig. 1: Reaction catalyzed by the SopB protein.
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): -- Gel Retardation Assay Structure Available (PDB or Homology model) -- PDB #: 4DID
Current Inhibitors: Expression Information (has it been expressed in bacterial cells): The plasmid pXX523 have been expressed in E. coli strains P678-54, DH1, HB101, and KY7231. pBR322 was used as the vector. Purification Method: http://www.jbc.org/content/264/26/15535.full.pdf+html Image of protein (PyMol with features delineated and shown separately):
Fig. 2: 4DID protein shown as cartoon. The GDP ligand is shown as sticks with magenta as carbon, blue as nitrogen, red as oxygen, and orange as phosphate.
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
*length of your protein in Amino Acids: 152 residues Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 16.756kiloDaltons Molar Extinction coefficient of your protein at 280 nm: 13980 M-1 cm-1
*GC% Content for gene: *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): *GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now) Primer design results for pNIC-Bsa4 cloning (list sequences of all of your ~40 nt long primers): (link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol) -- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
*NCBI Gene # or RefSeq#: NC_003197.1
*Protein ID (NP or XP #) or Wolbachia#: 4DID -- (need the NP or XP number here - put the PDB number down further 082413 - Dr. B)
*Organism (including strain): Salmonella enterica subsp. enterica serovar Typhimurium
Etiologic Risk Group (see link below): group 2
*Background/Disease Information (sort of like the Intro to your Mini Research Write up): The salmonella enterica serovar Typhi organism is responsible for typhoid fever. The bacteria works by invading the surface of the intestine, and spreading to deeper tissues (including that of the liver, spleen, and bone marrow). It causes 22 million cases of typhoid around the world and 200,000 deaths. It is most common in underdeveloped areas that do not have proper sanitation techniques, as it is transmitted through drinking water or eating food that is contaminated with the feces of an infected person.
Essentiality of this protein: SopB is an effector protein with phosphoinositide phosphatase activity. It has a distinct GTPase binding domain that interacts with the host Cdc42 cell to regulate critical events in eukaryotic cytoskeleton organization and membrane trafficking. The SopB protein mimics a host guanine nucleotide dissociation inhibitor both structurally and functionally by contacting key residues in the regulatory regions of Cdc42, which slows nucleotide exchange in the Cdc42 cell. It also prevents lysosomal degradation of the bacteria
Complex of proteins: May be complexed with the host Cdc42 cell.
Druggable Target: http://tdrtargets.org/drugs/view?mol_id=558832
*EC#: 3.1.3.36
Link to BRENDA EC# page:
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
-- Gel Retardation Assay
Structure Available (PDB or Homology model)
-- PDB #: 4DID
Current Inhibitors:
Expression Information (has it been expressed in bacterial cells): The plasmid pXX523 have been expressed in E. coli strains P678-54, DH1, HB101, and KY7231. pBR322 was used as the vector.
Purification Method: http://www.jbc.org/content/264/26/15535.full.pdf+html
Image of protein (PyMol with features delineated and shown separately):
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
*length of your protein in Amino Acids: 152 residues
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 16.756kiloDaltons
Molar Extinction coefficient of your protein at 280 nm: 13980 M-1 cm-1
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
*CDS Gene Sequence (paste as text only):
*GC% Content for gene:
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only):
*GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list sequences of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
Sources:
http://microbes.ucsc.edu/cgi-bin/hgGateway?db=salmTyph
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3339929/
http://www.jbc.org/content/264/26/15535.full.pdf+html (expression and purification)
http://tdrtargets.org/drugs/view?mol_id=558832
http://www.ncbi.nlm.nih.gov/nuccore/NC_003197.1?report=fasta&from=1178231&to=1179916&strand=true (gene sequence)
http://www.ncbi.nlm.nih.gov/gene/1252609
http://oba.od.nih.gov/oba/rac/guidelines/nih_guidelines.htm (risk group)